CpxR-Dependent Thermoregulation of Serratia marcescens PrtA Metalloprotease Expression and Its Contribution to Bacterial Biofilm Formation

Abstract

PrtA is the major secreted metalloprotease of Previous reports implicate PrtA in the pathogenic capacity of this bacterium. PrtA is also clinically used as a potent analgesic and anti-inflammatory drug, and its catalytic properties attract industrial interest. Comparatively, there is scarce knowledge about the mechanisms that physiologically govern PrtA expression in In this work, we demonstrate that PrtA production is derepressed when the bacterial growth temperature decreases from 37°C to 30°C. We show that this thermoregulation occurs at the transcriptional level. We determined that upstream of , there is a conserved motif that is directly recognized by the CpxR transcriptional regulator. This feature is found along strains irrespective of their isolation source, suggesting an evolutionary conservation of CpxR-dependent regulation of PrtA expression. We found that in s, the CpxAR system is more active at 37°C than at 30°C. In good agreement with these results, in a mutant background, is derepressed at 37°C, while overexpression of the NlpE lipoprotein, a well-known CpxAR-inducing condition, inhibits PrtA expression, suggesting that the levels of the activated form of CpxR are increased at 37°C over those at 30°C. In addition, we establish that PrtA is involved in the ability of to develop biofilm. In accordance, CpxR influences the biofilm phenotype only when bacteria are grown at 37°C. In sum, our findings shed light on regulatory mechanisms that fine-tune PrtA expression and reveal a novel role for PrtA in the lifestyle of We demonstrate that metalloprotease PrtA expression is transcriptionally thermoregulated. While strongly activated below 30°C, its expression is downregulated at 37°C. We found that in , the CpxAR signal transduction system, which responds to envelope stress and bacterial surface adhesion, is activated at 37°C and able to downregulate PrtA expression by direct interaction of CpxR with a binding motif located upstream of the gene. Moreover, we reveal that PrtA expression favors the ability of to develop biofilm, irrespective of the bacterial growth temperature. In this context, thermoregulation along with a highly conserved CpxR-dependent modulation mechanism gives clues about the relevance of PrtA as a factor implicated in the persistence of on abiotic surfaces and in bacterial host colonization capacity.

A CpxR-Regulated zapD Gene Involved in Biofilm Formation of Uropathogenic Proteus mirabilis

“…In contrast, E. coli CpxR-P can directly bind to the fimA promoter, thus negatively affecting the production of type 1 fimbriae and biofilms (44). S. marcescens CpxR negatively regulates biofilm formation by directly inhibiting the expression of PrtA (31). For the first time, we found that the P. mirabilis CpxR-regulated zapABCD genes (encoding T1SS and ZapA protease) participate in biofilm formation, which corresponds with the role of CpxAR TCS in the regulation of virulence factors.…”

“…4). Because we observed that ZapA expression was not associated with P. mirabilis autolysis (data not shown) as was shown to be the case for PrtA metalloprotease in S. marcescens (31), we can rule out the possibility that ZapA-dependent liberation of eDNA could act to modulate the formation or maintenance of P. mirabilis biofilms. Further studies are needed to reveal the scenarios of zapA involvement in the biofilm-forming process.…”

“…Aggregation might permit more adherence with bacteria piling on top of each other. It was suggested that a metalloprotease may modulate biofilm formation dependent on the catalytic activity or structural property harbored by it (31). Accordingly, we noticed that biofilm-deficient zapD and cpxR mutants had lower extracellular protease activity, production of eDNA and EPS, adhesion to the uroepithelial cells, and autoaggregative ability than those of the wild type (Fig.…”

“…Moreover, a metalloprotease was shown to contribute to biofilm formation in Serratia spp. (31). Therefore, we constructed the zapA mutant exhibiting no growth defect compared to the wild-type and zapA-complemented (zapAc) strains to evaluate the biofilm formation ability.…”

Comparative transcriptome analysis reveals metabolic regulation of prodigiosin in Serratia marcescens

“…Liu et al obtained a super PG-producer S. marcescens jx1 which could produce 6.5 g/L of PG by microwave mutagenesis technology [11]. Furthermore, several reports have shown the influence of temperature on the phenotypes or enzymes of S. marcescens, such as the temperature-dependent resistance of S. marcescens to norfloxacin (hydrophilic fluoroquinolone) and nalidixic acid (hydrophobic quinolone) [12], its swimming or swarming motility [13,14], its outer membrane vesicles [15], and its metalloproteases and serine proteases [16,17].…”